During chronic human immunodeficiency virus (HIV) infection, HIV-specific T cells fail to control viral replication, in part because they lose the ability to proliferate, produce cytokines, and lyse infected target cells. Functional restoration of HIV-specific T cells is therefore essential for limiting disease progression, and could also be used in combination with therapeutic vaccination to increase the functionality of vaccine generated HIV-specific T cell responses. Most attempts to restore activity in dysfunctional HIV-specific T cells by blocking or stimulating cell surface markers have not been successful. Recently, however, blocking signaling of death protein 1 (PD-1), an inhibitory receptor in the CD28 family, was shown to dramatically increase the frequency and function of virus-specific CD8+ T cells both in subjects with chronic HIV infection and in mice infected with lymphocytic choriomeningitis virus (LCMV), a model system for T cell dysfunction induced by chronic viral infection. Our preliminary data demonstrate that PD-1 is also specifically up-regulated on HIV-specific CD4+ T cells during chronic HIV infection and that blockade increases HIV-specific CD4+ T cell proliferation. However little is known about the effect of PD-1 blockade on HIV-specific CD4+ T cell cytokine production and maturation. PD-1 pathway blockage did result in a substantial decrease in LCMV viral load in chronically infected mice, but since there is no validated mouse model for chronic HIV infection, it has been difficult to determine whether the same would be true for HIV infection. We propose to use a two pronged in vitro and in vivo approach to determine if blockade of PD-1 ligation is a viable therapeutic approach for restoring or augmenting HIV-specific CD4+ T cell function in subjects with chronic HIV infection. We hypothesize that blockade of PD-1 binding in HIV-infected subject's peripheral blood mononuclear cells (PBMC) will specifically enhance HIV-specific CD4+ T cell function in vitro and that "cocktails" of mAbs which simultaneously block PD- 1 ligation and activate costimulatory molecules CD28, 4-1BB or OX40 will lead to even greater enhancement of function than PD-1 blockade alone. In addition we will examine the effects of blockade of PD-1 ligation on HIV viral load and CD4 count in vivo using a novel humanized Rag2-/-?c/- mouse (RAG-hu) model of HIV infection. Rag2-/-3c/- mice are reconstituted with human CD34 hematopoietic stems cells which differentiate into human immune cells capable of mounting immune responses. We have recently demonstrated that these mice are permissive to HIV infection. We hypothesize that blockade of PD-1 binding in HIV-infected RAG-hu mice will enhance control of HIV replication resulting in lower HIV viral loads and higher CD4+ T cell counts in vivo. The specific aims of this study are: 1)To enhance HIV-specific CD4+ T cell function in PBMC from chronically HIV-infected subjects by blocking the PD-1 pathway using anti PD-1 ligand mAbs and soluble PD-1. 2) To validate the RAG-hu mouse model of chronic HIV infection for the study of HIV-specific T cell responses and use it to investigate the effects of blocking the PD-1 pathway on HIV replication in vivo. PUBLIC HEALTH RELEVANCE: The human immunodeficiency virus (HIV) is estimated to have infected 40 million people and to be responsible for 2.9 million deaths worldwide in 2006. Development of immune-based therapies to increase the function of HIV-specific CD4+ T cells are vital for controlling the disease. PD-1 pathway blocking therapies could slow disease progression and be used in combination with therapeutic vaccination to increase the functionality of vaccine generated HIV-specific CD4+ T cell responses.